About this Author

College chemistry, 1983

The 2002 Model

After 10 years of blogging. . .

Derek Lowe, an Arkansan by birth, got his BA from Hendrix College and his PhD in organic chemistry from Duke before spending time in Germany on a Humboldt Fellowship on his post-doc. He's worked for several major pharmaceutical companies since 1989 on drug discovery projects against schizophrenia, Alzheimer's, diabetes, osteoporosis and other diseases.
To contact Derek email him directly: derekb.lowe@gmail.com
Twitter: Dereklowe

March 15, 2006

No Immunity From Immunity

There's been a severe problem with the first-in-man dosing of a new antibody in England. TeGenero, a small company in Germany, was testing their most advanced candidate, TGN1412, which is a monoclonal antibody against CD28. That's a glycoprotein on the surface of many types of T cells, and it seems to be extremely important in several mechanisms that activate them. Accordingly, finding something to modulate its activity would seem to be a promising way to attack autoimmune and other inflammatory diseases.

But when TGN1412 went into six volunteers in Phase I this week, all of them ended up in the hospital, and two of them are gravely ill. The press reports make it seem like some sort of anaphylactic reaction, but it sounds like they're not treating it like simple anaphylaxsis, so there must be something more going on. That's a very surprising outcome, since if anything you'd expect the immune response to be downregulated in response to the drug. (Update: this is only partly true - see end of post.) But there's an awful lot about human immunology that we don't understand, to put it mildly. TeGenero says that:

"These events were completely unexpected and do not reflect the results we obtained from initial laboratory studies which enabled us to progress investigations into human volunteers. . .The drug was developed in accordance with all regulatory and clinical guidelines and standards. In pre-clinical studies, TGN1412 has been shown to be safe and the reactions which occurred in these volunteers were completely unexpected”

I have to believe that this is correct. Neither TeGenero, the company conducting the trial for them (Parexel), or the British regulatory authorities have anything to gain from taking a drug into humans that had shown severe effects in animals, despite rumors to the contrary that seem to be going around in England. There's a breathless report in The Independent (I know, that's redundant) to this effect:

"The men, who were offered £2,000 to take part, were recruited by the US company Parexel, for the trial in its 36-bed unit on the Northwick Park hospital campus. They reportedly signed a contract warning that side-effects in rats and mice included "increased urine volume, decreased faeces, redness of the skin". Dogs experienced "increased heart rate and decreased blood pressure"."

I have news for the Independent, though - if we killed off every drug that showed effects like this, we'd never take anything into humans at all. The dog cardiovascular effects would be worth checking out in detail before going forward, naturally, but that's what dogs are for, to show you cardiovascular side effects. And I note that none of these effects have anything to do with devastating immune reactions. If that's all that TeGenero saw in the animal studies, I can see why they were shocked at the human results.

But antibodies are powerful - no one should forget that. Every antibody therapy has a small but real risk of setting off something terrible, and if you're targeting the immune response itself, well, the risk just goes up. The immune system is a bit like demonlogy: don't call anything up that you don't know how to send back down. Unfortunately, we don't know many effective spells yet.

My sympathies go out to those affected and their relatives and friends, of course. They shouldn't, as far as I can see, torture themselves thinking that this could have been foreseen. I hope that everyone makes it through.

Update: I've inadvertently glossed over some of the mechanism of TGN1412. Its binding to CD28 actually sets off the receptor's signaling - it's an antibody agonist. This makes it a T-cell activator, but it seems to particularly activate the class known as regulatory T-cells. These are modulators of the activity of other classes of T-cells involved in autoimmune responses.

So there's a way that things could have gone wrong - if TGN1412 isn't quite as selective in the real-world human immune system as it is in the animal models. Alternatively, even if it is selective but the spectrum of human T-cell response to CD28 ligands is intrinsically somewhat different, things could have gone off the rails very quickly. There appears to have been little reason to suspect either of these possibilities going into the clinic. We're going to learn something important about human immunology from this incident, but this is certainly a hell of a way to do it.

I believe that TGN1412 is actually an agonist antibody to CD28, and was being developed to increase immmune response against cancers by activating T cells. It's easier to understand the symptomology that way, as well.

This is a really tragic case, the worst I've heard of with any biologic. I hope everyone recovers.

This is a terrible thing to happen to the trial volunteers. The accounts of what happened sound horrific, and unfortunately two of the patients remain critically ill.

Last night, the BBC news said that two other possibilities could be a problem with drug purity or an accidental high dose given to the patients. I'm not involved in this particular area, but are any of these more likely explanations than a reaction to the antibody treatment?

The press stories remain confused, with some claiming that the dosage was adjusted to the weight of the participants (normal for antibody dosing?) and that the two volunteers on life support machines received the highest dose.

Assuming that the intended dose was given, it is possible that the antibody was a much more effective immunostimulant in vivo in man than in animal or ex vivo studies. Another possibility would be contamination of the batch of antibody employed in the study with e.g. LPS.

TGN1412 creates a hyper-response by NK cells. The immune system's safeguard against rampant auto-immunity is triggering cell death of the excess NK-cells through apoptosis.

Immuno-supressive drugs, such as Cyclosporin, you think should be used to treat an "anaphylactic reaction" make NK cells immune to the apoptosis signal, thereby preventing a down-regulation of the reaction you are trying to halt.

Is it possible these men suffered "cytokine storm", that for some reason did not show itself in animal testing?

There's a possible clue on the manufacturer's own website:

"A pronounced T-cell activation and expansion mediated by CD28-SuperMABÂ® in animal models is accompanied by the expression of anti-inflammatory cytokines, like IL-10, rather than by the toxic cytokine storm of pro-inflammatory mediators induced by other agents that address the TCR complex."

If you have 8 people, 2 getting placebo, 4 who are in bad shape and 2 who are in really bad shape, that looks a lot like a dose-response curve. It does make you wonder why they gave them all more-or-less simultaneously.

It'd also be useful to see what the results from the primate studies were. Surely they ran some...

We can't really conclude much about what happened until we know whether the doses received by the volunteers were both correct and uncontaminated.
When you think of all the steps involved it's easy to conceive of a catastrophic screwup causing problems like this, though thst's the same reason why so many checks are needed.
Per a previous I don't think that 1st-in-human doses are usually determined by weight, but if someone prepared a dose in mg as a dose in mg/kg you get about 100 x the intended dose.
And don't think 100 kg is an unusual weight for a volunteer study. Given that so many Phase 1 trials are run in the UK an industry joke is that they don't tell you how the drug behaves in humans, just in humans who happen to be white males in their 20s who happen to be unemployed Rugby players.
I see how something like LPS contamination could produce effects like those seen, but I'm guessing LPS would be on the short list of stuff to screen for in any mfr'ing process. Especially at concentrations sufficient to produce such strong reactions. Could it be really, really bad saline? A synergistic reaction between low level contaminants and the antibody being studied?

There's a paper showing that the antibody worked on human T cells to stimulate the CD28-dependent pathway. But the problem is that there's no way to know what other T cell types are affected, or how molecules secreted by those T cells would interact with the rest of the immune system.

There are just a few sentences like the one quoted. The data in the paper is mostly for mice and rats. I'm not an expert in this area; I just scanned the references on the TeGenaro website, and did a PubMed on CD28.

So there's a way that things could have gone wrong - if TGN1412 isn't quite as selective in the real-world human immune system as it is in the animal models.

Surely the opposite is more likely and equally dangerous? It's an antibody drug, raised against the human protein. Thus you'd expect it to have highest effect on the human protein. In an animal model, the differences may mean it no longer binds to its main target as well, leading to a changed balance between on- and off-target effects.

And isn't that what we see? It was designed as an inflammatory activator. They then test it in animals, find that it activates regulatory T-cells as well, leading to a net suppression of inflammation. Put it back in humans, and the orignal designed activity comes back, giving huge inflammatory activation.

I carefully checked the literature on CD28 superagonists (this is how the Mab is called) on PubMed. Most of the work has been done by the groups of Thomas Hünig at the University of Würzburg. I could not find a single paper where they look at the effect on human cells in vitro. There is review (the one mentioned by Jim) that mentions an effect on human cells, but the effect is not discussed. There are strong data showing an induction of rat Tregs in vitro by the superagonist, but I have not been able to find a single experiment where they did the same using human cells, which is quite easy to do once you ahve the rat system going.
What this superagonist does is that it ligates many CD28 molecules together, therefore overcoming the need for a second stimulus to induce T cell activation.

A bit naive to use a "superstimulus" and not to expect a disaster. Anti CD28 will not only stimulate T-cells but also trigger the eosinophils to release Neurotixins etc. Biological systems are based on heterogeneity and regulated acordingly by using "logical AND gating" to avoid disregulation. There seems to be the believe that CD28 only exists on T-cells. But that probably comes because a lot of people only look at those receptors on T-cells

What also worries me is the idea to inject all volunteers at the same time. So if something goes acutely wrong you definitively have no chance. But perhaps we learn a lesson.

Newbie to this blog, just wanted to add that there was one study using the (very new) humanized mouse model (see below). I have to say I'm surprised that no journalists seem to have yet got hold of a copy of the phase I protocol, which surely must describe the safety experiments they did in animals (including non human primates).

One slight twist to this story is that there remains some controversy over what "regulatory" T cells are exacttly; some of the markers used to categorize this population may only be expressed transiently in vivo, plus they have to be artifically activated in vitro in order to exert their suppressive activity (and no one really knows precisely how this works in vivo). For a perspective on this, see Brigitta Stockingers review "T cell regulation: a special job or everyone's responsibility?" which is available free online here:

One last aside: it appears that the most severely affected individual in the study may have been the youngest, suggesting that activation of naive T cells (which are most abundant in youth and decline with age) may have contributed to the terrible outcome of this study.

Department of Cell Biology and Histology, Academic Medical Center of the University of Amsterdam (AMC-UvA), Amsterdam, The Netherlands.

Efficient and quick reconstitution of T cell compartments in lymphopenic patients is of great importance to prevent opportunistic infections, but remains difficult to achieve. Human T cell proliferation in a TCR-independent manner is possible in vitro with superagonist anti-CD28 antibodies, and such molecules are therefore promising therapeutic tools. Here, we investigated the in vivo effects of superagonist anti-CD28 treatment on human developing and mature T cells, in the recently developed model of "human immune system" BALB/c Rag2(-/-)gammac(-/-) mice. Our results show that superagonist anti-CD28 treatment transiently induces a 7-fold increase in thymocytes numbers and up to 18-fold accumulation of mature thymocytes. The increased thymic production lead to transient accumulation of mature T cells in the periphery at the peak of treatment effect (day 6). In addition, long-term peripheral T cell depletion was induced. Furthermore, the concomitant selective expansion and accumulation of suppressive CD4(+)CD25(+)FoxP3(+) T cells was induced in a transient manner. Superagonist anti-CD28 therapy could therefore be of clinical interest in humans, both for beneficial effect on thymic T cell production as well as regulatory T cell accumulation.

TeGenero AG announced today that its Humanized Agonistic Anti-CD28 Monoclonal Antibody TGN1412, which is currently in late-stage preclinical development, has received designation as orphan medicinal product from the European Medicines Agency (EMEA) for the treatment of B-cell Chronic Lymphocytic Leukaemia, B-CLL.
The decision of this positive opinion was announced after the March meeting of the Committee for Orphan Medicinal Products.
B-CLL is a rare, chronic and life-threatening leukaemic disease for which currently no curative treatment exists. The pathophysiology of B-CLL is closely linked to a pronounced dysfunction of the patients’ T lymphocytes. According to the proposed therapeutic mode-ofaction, enhanced activation of T lymphocytes by treatment with TeGenero’s TGN1412 is expected to result in fewer infections and better control of the cancer.
“The designation reflects a high medical need for safe and efficacious new treatment options in B-CLL”, says Dr. Thomas Hanke, Chief Scientific Officer of TeGenero. “Due to its novel mode-of-action, TGN1412 has the potential to provide a significant benefit to patients suffering from B-CLL, as demonstrated in recent pre-clinical studies.”
The EMEA orphan drug designation entitles TeGenero AG to exclusive marketing rights in the EU on TGN1412 for ten years following marketing approval and to protocol assistance by EMEA in order to optimize TeGenero´s drug development strategy in compliance with regulatory requirements.
About designation as orphan medicinal product
“Orphan drugs“ are medicinal products used for rare, life-threatening diseases or chronically debilitating conditions where no other or no sufficient effective treatment exists. Benefits of designation as orphan medicinal product by the EMEA include reduced fees for centralized activities as well as advice on the conduct of clinical trials. An orphan designation is not a marketing authorization, which can only be granted after the quality, safety and efficacy of the product have been demonstrated.

About TGN1412
TGN1412 is a Humanized Agonistic Anti-CD28 Monoclonal Antibody that is being developed by TeGenero AG for the treatment of autoimmune and oncological diseases including rheumatoid arthritis and B-CLL. It is in late-stage preclinical development with clinical trials anticipated to start in Q4, 2005.

About TeGenero AG:
TeGenero is a biopharmaceutical company dedicated to the identification and development of innovative, highly effective and broadly applicable immunotherapeutics, thus providing patients the means to regain a functional and balanced immune system.
The company was founded in 2000 by Prof. Thomas Hünig, member of the supervisory board, together with scientific co-workers of the University of Würzburg. The supervisory board is chaired by Prof. Jürgen Drews and includes Prof. Patrick Baeuerle of Micromet AG. TeGenero’s management consists of Dr. Benedikte Hatz, CEO, and Dr. Thomas Hanke, CSO.
The headquartars is in the BioMed Innovation and Founding Center at the Science-Park Würzburg.

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